The functional role of the cell membrane in the regulation of transmembrane and trans-epithelial water and electrolyte transport remains one of the most challenging and fundamental problems in cell biology. It is now generally agreed that such regulation is mediated, at least in part, by ouabain-sensitive Na-K-ATPase although the subcellular location of this enzyme in structurally complex osmoregulatory tissues is not understood well. The objectives of the proposed research are to explore the closely linked functional relationships between membrane specialization, sodium transport sites, Na-K-ATPase activity and the routes of transepithelial electrolyte transport across reabsorptive and secretory epithelia. These will be studied by: (1) high resolution electron microscopy, (2) Na-K-ATPase cytochemistry at the ultrastructural level, and (3) autoradiographic analysis of tritiated-ouabain binding sites. In addition, we will examine by physiological methods the relationship between the localization of ouabain-sensitive sodium pump sites in a model secretory system, the avian salt gland, and ouabain-sensitive and insensitive ion fluxes in methacholine-stimulated and non-stimulated slices as they are revealed by established tracer techniques. We anticipate that such correlated physiological and cytochemical studies will help to elucidate the specific role of Na-K-ATPase-dependent and independent sodium pump sites in the mechanism of ion secretion. Moreover, in addition to obtaining critical data relating to the over-all problem of electrolyte transport in reabsorptive and secretory epithelia, we expect that the methodology which will be developed and refined during the course of these studies will have wide appplicability to related problems in other tissues.